Heterocyclic compounds as kinase inhibitors

ABSTRACT

The present invention is directed to certain amides and heterocyclic compounds. The present invention also relates to uses of these compounds to treat several diseases including autoimmune disorders, cardiovascular disorders, inflammation, central nervous system disorders, arterial thrombotic disorders, fibrotic disorders, glaucoma, and neoplastic disorders.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a continuation of U.S. patent application Ser. No.15/574,800, filed Nov. 16, 2017 (published as US20180141959), which isthe U.S. National Stage of International Application No.PCT/US2016/033111, filed May 18, 2016, which claims priority to U.S.Provisional Patent Application No. 62/203,070, filed Aug. 10, 2015, andU.S. Provisional Patent Application No. 62/163,369, filed May 18, 2015,the contents of each of which are incorporated herein by reference intheir entireties.

FIELD OF THE INVENTION

The present invention relates to heterocyclic compounds, theircompositions and medicaments containing the same, as well as processesfor the preparation and use of such compounds, compositions andmedicaments. Such compounds are potentially useful in the treatment ofdiseases associated with inappropriate tyrosine and/or serine/threoninekinase activity.

BACKGROUND OF THE INVENTION

An important large family of enzymes is the protein kinase enzymefamily. Currently, there are about 500 different known protein kinases.Protein kinases serve to catalyze the phosphorylation of an amino acidside chain in various proteins by the transfer of the γ-phosphate of theATP-Mg²⁺ complex to said amino acid side chain. These enzymes controlthe majority of the signaling processes inside cells, thereby governingcell function, growth, differentiation and destruction (apoptosis)through reversible phosphorylation of the hydroxyl groups of serine,threonine and tyrosine residues in proteins. Studies have shown thatprotein kinases are key regulators of many cell functions, includingsignal transduction, transcriptional regulation, cell motility, and celldivision. Several oncogenes have also been shown to encode proteinkinases, suggesting that kinases play a role in oncogenesis. Theseprocesses are highly regulated, often by complex intermeshed pathwayswhere each kinase will itself be regulated by one or more kinases.Consequently, aberrant or inappropriate protein kinase activity cancontribute to the rise of disease states associated with such aberrantkinase activity. Due to their physiological relevance, variety andubiquitousness, protein kinases have become one of the most importantand widely studied family of enzymes in biochemical and medicalresearch.

The protein kinase family of enzymes is typically classified into twomain subfamilies: Protein Tyrosine Kinases and Protein Serine/ThreonineKinases, based on the amino acid residue they phosphorylate. Theserine/threonine kinases (PSTK), includes cyclic AMP- and cyclicGMP-dependent protein kinases, calcium- and phospholipid-dependentprotein kinase, calcium- and calmodulin-dependent protein kinases,casein kinases, cell division cycle protein kinases and others. Thesekinases are usually cytoplasmic or associated with the particulatefractions of cells, possibly by anchoring proteins. Aberrant proteinserine/threonine kinase activity has been implicated or is suspected ina number of pathologies such as rheumatoid arthritis, psoriasis, septicshock, bone loss, many cancers and other proliferative diseases.Accordingly, serine/threonine kinases and the signal transductionpathways which they are part of are important targets for drug design.The tyrosine kinases phosphorylate tyrosine residues. Tyrosine kinasesplay an equally important role in cell regulation. These kinases includeseveral receptors for molecules such as growth factors and hormones,including epidermal growth factor receptor, insulin receptor, plateletderived growth factor receptor and others. Studies have indicated thatmany tyrosine kinases are transmembrane proteins with their receptordomains located on the outside of the cell and their kinase 5 domains onthe inside. Much work is also under progress to identify modulators oftyrosine kinases as well.

A major signal transduction systems utilized by cells is theRhoA-signaling pathways. RhoA is a small GTP binding protein that can beactivated by several extracellular stimuli such as growth factor,hormones, mechanic stress, osmotic change as well as high concentrationof metabolite like glucose. RhoA activation involves GTP binding,conformation alteration, post-translational modification(geranylgeranyllization and famesylation) and activation of itsintrinsic GTPase activity. Activated RhoA is capable of interacting withseveral effector proteins including ROCKs and transmit signals intocellular cytoplasm and nucleus.

ROCK1 and 2 constitute a family of kinases that can be activated byRhoA-GTP complex via physical association. Activated ROCKs phosphorylatea number of substrates and play important roles in pivotal cellularfunctions. The substrates for ROCKs include myosin binding subunit ofmyosin light chain phosphatase (MBS, also named MYPT1), adducin, moesin,myosin light chain (MLC), LIM kinase as well as transcription factorFHL. The phosphorylation of theses substrates modulate the biologicalactivity of the proteins and thus provide a means to alter cell'sresponse to external stimuli. One well documented example is theparticipation of ROCK in smooth muscle contraction. Upon stimulation byphenylephrine, smooth muscle from blood vessels contracts. Studies haveshown that phenylephrine stimulates alpha-adrenergic receptors and leadsto the activation of RhoA. Activated RhoA in turn stimulates kinaseactivity of ROCK1 and which in turn phosphorylates MBS. Suchphosphorylation inhibits the enzyme activity of myosin light chainphosphatase and increases the phosphorylation of myosin light chainitself by a calcium-dependent myosin light chain kinase (MLCK) andconsequently increases the contractility of myosin-actin bundle, leadingto smooth muscle contraction. This phenomena is also sometimes calledcalcium sensitization. In addition to smooth muscle contraction, ROCKshave also been shown to be involved in cellular functions includingapoptosis, cell migration, transcriptional activation, fibrosis,cytokinesis, inflammation and cell proliferation. Moreover, in neuronsROCK plays a critical role in the inhibition of axonal growth bymyelin-associated inhibitory factors such as i myelin-associatedglycoprotein (MAG). ROCK-activity also mediates the collapse of growthcones in developing neurons. Both processes are thought to be mediatedby ROCK-induced phosphorylation of substrates such as LIM kinase andmyosin light chain phosphatase, resulting in increased contractility ofthe neuronal actin-myosin system. Inhibitors of ROCKs have beensuggested for use in the treatments of a variety of diseases. Theyinclude cardiovascular diseases such as hypertension, chronic andcongestive heart failure, cardiac hypertrophy, restenosis, chronic renalfailure and atherosclerosis. In addition, because of its muscle relaxingproperties, it is also suitable for asthma, male erectile dysfunctions,female sexual dysfunction and over-active bladder syndrome. ROCKinhibitors have been shown to possess anti-inflammatory properties. Thusthey can be used as treatment for neuroinflammatory diseases such asstroke, multiple sclerosis, Alzheimer's disease, Parkinson's disease,amyotrophic lateral sclerosis and inflammatory pain, as well as otherinflammatory diseases such as rheumatoid arthritis, irritable bowelsyndrome, inflammatory bowel disease. In addition, based on theirneurite outgrowth inducing effects, ROCK inhibitors could be usefuldrugs for neuronal regeneration, inducing new axonal growth and axonalrewiring across lesions within the CNS. ROCK inhibitors are thereforelikely to be useful for regenerative (recovery) treatment of CNSdisorders such as spinal cord injury, acute neuronal injury (stroke,traumatic brain injury), Parkinson's disease, Alzheimer's disease andother neurodegenerative disorders.

Since ROCK inhibitors reduce cell proliferation and cell migration, theycould be useful in treating cancer and tumor metastasis. Furthermore,there is evidence suggesting that ROCK inhibitors suppress cytoskeletalrearrangement upon virus invasion, thus they also have potentialtherapeutic value in antiviral and anti-bacterial applications. ROCKinhibitors may also be useful for the treatment of insulin resistanceand diabetes.

The inventors have discovered novel heterocyclic compounds, which areinhibitors of ROCK activity. Such derivatives are useful in thetreatment of disorders associated with inappropriate ROCK activity.

SUMMARY OF THE INVENTION

The present invention is directed to a compound of Formula I:

or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof;

or a pharmaceutically acceptable salt, solvate, hydrate orphysiologically functional derivative thereof;

A is indazol-3-yl, pyrazol-4-yl,

wherein (i) G is CR′ or N; (ii) X is hydrogen, C₁₋₆ alkyl, C₃₋₇cycloalkyl, —OR₂ or —NR₃R₄; and (iii) R′, R″, R₂, R₃ and R₄ areindependently —H or C₁₋₆ alkyl or, C₃₋₇ cycloalkyl;

Z is selected from the group consisting of:

wherein (i) R₅ is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl; (ii) R₇ and R₈ areindependently —H, halo, C₁₋₆ alkyl, C₃₋₇ cycloalkyl, —O—(C₁₋₆ alkyl),—OH, —CN, —COOR′, —OC(O)R′, NHR′, N(R′)₂, —NHC(O)R′, —NHS(O)₂R′,—C(O)NHR′, or —S(O)₂R′ wherein R′ is —H, C₁₋₆ alkyl, or C₃₋₇ cycloalkyl;(iii) G₁, G₂ and G₃ are independently CH or N.

Z′ is a bond, O or NR₆, wherein R₆ is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl;

R is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl;

R₁ is —H or C₁₋₆ alkyl;

Q is a bond or C₁₋₆ alkyl;

J is a bond or C₁₋₆ alkyl;

W is —H, —OR₉, —NR₁₀R₁₁, or —S(O)_(m)R₁₂, wherein (i) R₉, R₁₀ and R₁₁are independently —H, C₁₋₆ alkyl, C₃₋₇ cycloalkyl, formyl, C₁₋₆alkylcarbonyl, C₃₋₇ cycloalkylcarbonyl, or C₁₋₆ alkylsulfonyl;

(ii) m is an integer from 0 to 2; and (iii) R₁₂ is C₁₋₆ alkyl or C₃₋₇cycloalkyl; and

Ar is a phenyl, naphthyl, or C₅₋₁₀ heterocycle, each of which isoptionally substituted with halo, —OH, —CN, —COOR_(a), —OR_(a), —SR_(a),—OC(O)R_(a), —NHR_(a), —NR_(a)R_(b), —NHC(O)R_(a), —NHC(O)NR_(a)R_(b),—C(O)NR_(a)R_(b), —NS(O)₂R_(a), —S(O)₂NR_(a)R_(b), —S(O)₂R_(a),guanidino, nitro, nitroso, C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl, or 3- to10-membered heterocycle, wherein the C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl,or 3 to 10-membered heterocycle is unsubstituted or substituted with oneor more of halo, —OH, —CN, —COOR_(a), —OR_(a), —SR_(a), —OC(O)R_(a),—NHR_(a), —NR_(a)R_(b), —NHC(O)R_(a), —NHC(O)NR_(a)R_(b),—C(O)NR_(a)R_(b), —NS(O)₂R_(a), —S(O)₂NR_(a)R_(b), —S(O)₂R_(a),guanidino, nitro, nitroso, C₁₋₆ alkyl, aryl, or C₃₋₇ cycloalkyl; whereineach of R_(a) and R_(b) is independently H or C₁₋₆ alkyl; and optionallyR_(a) and R_(b) together attaching to N or O form a 4- to 8-memberedheterocycle.

In one embodiment, the present invention is directed to a compound ofFormula II:

wherein (i) G is CR′ or N; (ii) X is hydrogen, C₁₋₆ alkyl, C₃₋₇cycloalkyl, —OR₂ or —NR₃R₄; and

(iii) R′, R″, R₂, R₃ and R₄ are independently —H or C₁₋₆ alkyl or, C₃₋₇cycloalkyl; Z is selected from the group consisting of:

wherein (i) R₅ is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl; (ii) R₇ and R₈ areindependently —H, halo, C₁₋₆ alkyl, C₃₋₇ cycloalkyl, —O—(C₁₋₆ alkyl),—OH, —CN, —COOR′, —OC(O)R′, NHR′, N(R′)₂, —NHC(O)R′, —NHS(O)₂R′,—C(O)NHR′, or —S(O)₂R′ wherein R′ is —H, C₁₋₆ alkyl, or C₃₋₇ cycloalkyl;(iii) G₂, G₃ and G₄ are independently CH or N.

Z′ is a bond, O or NR₆, wherein R₆ is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl;

R is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl;

R₁ is —H or C₁₋₆ alkyl;

Q is a bond or C₁₋₆ alkyl;

J is a bond or C₁₋₆ alkyl;

W is —H, —OR₉, —NR₁₀R₁₁, or —S(O)_(m)R₁₂, wherein (i) R₉, R₁₀ and R₁₁are independently —H, C₁₋₆ alkyl, C₃₋₇ cycloalkyl, formyl, C₁₋₆alkylcarbonyl, C₃₋₇ cycloalkylcarbonyl, or C₁₋₆ alkylsulfonyl;

-   -   (ii) m is an integer from 0 to 2; and (iii) R₁₂ is C₁₋₆ alkyl or        C₃₋₇ cycloalkyl; and

Ar is a phenyl, naphthyl, or C₅₋₁₀ heterocycle, each of which isoptionally substituted with halo, —OH, —CN, —COOR_(a), —OR_(a), —SR_(a),—OC(O)R_(a), —NHR_(a), —NR_(a)R_(b), —NHC(O)R_(a), —NHC(O)NR_(a)R_(b),—C(O)NR_(a)R_(b), —NS(O)₂R_(a), —S(O)₂NR_(a)R_(b), —S(O)₂R_(a),guanidino, nitro, nitroso, C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl, or 3- to10-membered heterocycle, wherein the C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl,or 3 to 10-membered heterocycle is unsubstituted or substituted with oneor more of halo, —OH, —CN, —COOR_(a), —OR_(a), —SR_(a), —OC(O)R_(a),—NHR_(a), —NR_(a)R_(b), —NHC(O)R_(a), —NHC(O)NR_(a)R_(b),—C(O)NR_(a)R_(b), —NS(O)₂R_(a), —S(O)₂NR_(a)R_(b), —S(O)₂R_(a),guanidino, nitro, nitroso, C₁₋₆ alkyl, aryl, or C₃₋₇ cycloalkyl; whereineach of R_(a) and R_(b) is independently H or C₁₋₆ alkyl; and optionallyR_(a) and R_(b) together attaching to N or O form a 4- to 8-memberedheterocycle.

In another embodiment, the present invention is directed to a compoundof Formula III:

Wherein A is indazol-3-yl, pyrazol-4-yl,

wherein (i) G is CR′ or N; (ii) X is hydrogen, C₁₋₆ alkyl, C₃₋₇cycloalkyl, —OR₂ or —NR₃R₄; and (iii) R′, R″, R₂, R₃ and R₄ areindependently —H or C₁₋₆ alkyl or, C₃₋₇ cycloalkyl;

G₅, G₆ and G₇ are independently CH or N.

Z′ is a bond, O or NR₆, wherein R₆ is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl;

R is —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl;

R₁ is —H or C₁₋₆ alkyl;

Q is a bond or C₁₋₆ alkyl;

J is a bond or C₁₋₆ alkyl;

W is —H, —OR₉, —NR₁₀R₁₁, or —S(O)_(m)R₁₂, wherein (i) R₉, R₁₀ and R₁₁are independently —H, C₁₋₆ alkyl, C₃₋₇ cycloalkyl, formyl, C₁₋₆alkylcarbonyl, C₃₋₇ cycloalkylcarbonyl, or C₁₋₆ alkylsulfonyl;

(ii) m is an integer from 0 to 2; and (iii) R₁₂ is C₁₋₆ alkyl or C₃₋₇cycloalkyl; and

Ar is a phenyl, naphthyl, or C₅₋₁₀ heterocycle, each of which isoptionally substituted with halo, —OH, —CN, —COOR_(a), —OR_(a), —SR_(a),—OC(O)R_(a), —NHR_(a), —NR_(a)R_(b), —NHC(O)R_(a), —NHC(O)NR_(a)R_(b),—C(O)NR_(a)R_(b), —NS(O)₂R_(a), —S(O)₂NR_(a)R_(b), —S(O)₂R_(a),guanidino, nitro, nitroso, C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl, or 3- to10-membered heterocycle, wherein the C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl,or 3 to 10-membered heterocycle is unsubstituted or substituted with oneor more of halo, —OH, —CN, —COOR_(a), —OR_(a), —SR_(a), —OC(O)R_(a),—NHR_(a), —NR_(a)R_(b), —NHC(O)R_(a), —NHC(O)NR_(a)R_(b),—C(O)NR_(a)R_(b), —NS(O)₂R_(a), —S(O)₂NR_(a)R_(b), —S(O)₂R_(a),guanidino, nitro, nitroso, C₁₋₆ alkyl, aryl, or C₃₋₇ cycloalkyl; whereineach of R_(a) and R_(b) is independently H or C₁₋₆ alkyl; and optionallyR_(a) and R_(b) together attaching to N or O form a 4- to 8-memberedheterocycle.

In some embodiments, the present invention is directed to a compound ofFormula I, II, and/or III selected from the group consisting of:

The present invention is also directed to a compound of Formula V:

or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof, or a pharmaceutically acceptable salt, solvate,hydrate or physiologically functional derivative thereof,

wherein:

A is indazol-3-yl, pyrazol-4-yl or

wherein (i) G is CH or N; and (ii) X is hydrogen, —OR₂ or —NR₃R₄,wherein each of R₂, R₃ and R₄ is independently —H or C₁₋₆ alkyl;

each of R₁₃ and R₁₄ is independently —H, halo, C₁₋₆ alkyl, or C₃₋₇cycloalkyl;

each of R₁₅ and R₂₀ is independently —H, halo, —OH, —CN, —COOR′, —OR′,—SR′, —OC(O)R′, —NHR′, —NR′R″, —NHC(O)R′, —NHC(O)NR′R″, —C(O)NR′R″,—NS(O)₂R′, —S(O)₂NR′R″, —S(O)₂R′, guanidino, nitro, nitroso, C₁₋₆ alkyl,aryl, C₃₋₇ cycloalkyl, and 3- to 10-membered heterocycle, wherein eachof the C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl, or 3- to 10-memberedheterocycle independently is unsubstituted or substituted with one ormore of halo, —OH, —CN, —COOR′, —OR′, —SR′, —OC(O)R′, —NHR′, —NR′R″,—NHC(O)R′, —NHC(O)NR′R″, —C(O)NR′R″, —NS(O)₂R′, —S(O)₂NR′R″, —S(O)₂R′,guanidino, nitro, nitroso, C₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl; whereineach of R′ and R″ is independently —H or C₁₋₆ alkyl; and optionally R′and R″ together attaching to N or O form a 4- to 8-membered heterocycle;

each of R₁₆, R₁₇, R₁₈ and R₁₉ is independently —H, C₁₋₆ alkyl, aryl,C₃₋₇ cycloalkyl, or 3 to 10-membered heterocycle; wherein the C₁₋₆alkyl, aryl, C₃₋₇ cycloalkyl, or 3- to 10-membered heterocycle isunsubstituted or substituted with one or more of halo, —OH, —CN,—COOR_(a), —OR_(a), —SR_(a), —OC(O)R_(a), —NHR_(a), —NR_(a)R_(b),—NHC(O)R_(a), —NHC(O)NR_(a)R_(b), —C(O)NR_(a)R_(b), —NS(O)₂R_(a),—S(O)₂NR_(a)R_(b), —S(O)₂R_(a), guanidino, nitro, nitroso, C₁₋₆ alkyl,aryl, C₃₋₇ cycloalkyl; wherein each of R_(a) and R_(b) is independently—H or C₁₋₆ alkyl; and optionally R_(a) and R_(b) together attaching to Nor O forms a 4- to 8-membered heterocycle; and

J is a bond or C₁₋₆ alkyl.

In certain aspects, A is pyrazol-4-yl. In other aspects, A ispyridine-4-yl. In one aspect, the present invention relates to acompound of Formula II wherein both R₁₃ and R₁₄ are methyl.

In some embodiments, the present invention is directed to a compound ofFormula V is selected from the group consisting of:

In certain aspects, the present invention provides a compound asdisclosed herein for use in treating a disease related to upregulationof Rho kinase-signaling pathways.

In other aspects, the present invention is directed to a method oftreating an autoimmune disorder in a subject comprising: administeringto the subject a therapeutically effective amount of a compounddisclosed herein. In one aspect, the autoimmune disorder is rheumatoidarthritis, multiple sclerosis, systemic lupus erythematosus (SLE),psoriasis, Crohn's disease, atopic dermatitis, eczema, or graft-versus-host disease (GVHD).

In some embodiments, the present invention provides a method of treatinga cardiovascular disorder in a subject comprising: administering to thesubject a therapeutically effective amount of a compound disclosedherein. In one embodiment, the cardiovascular disorder is hypertension,atherosclerosis, restenosis, cardiac hypertrophy, ocular hypertension,cerebral ischemia, cerebral vasospasm, or erectile dysfunction.

In other embodiments, the present invention provides a method oftreating inflammation in a subject comprising: administering to thesubject a therapeutically effective amount of a compound disclosedherein. In certain aspects, the inflammation is asthma, cardiovascularinflammation, renal inflammation or arteriosclerosis.

In certain aspects, the present invention provides a method of treatinga central nervous system disorder in a subject comprising: administeringto the subject a therapeutically effective amount of a compounddisclosed herein. In one aspect, the central nervous system disorder isneuronal degeneration or spinal cord injury. In another aspect, thecentral nervous system disorder is Huntington's disease, Parkinson'sDisease, Alzheimer's, Amyotrophic lateral sclerosis (ALS), or multiplesclerosis.

The present invention also provides a method of treating an arterialthrombotic disorder in a subject comprising: administering to thesubject a therapeutically effective amount of a compound disclosedherein. In one embodiment, the arterial thrombotic disorder is plateletaggregation, or leukocyte aggregation.

In other aspects, the present invention relates to a method of treatinga fibrotic disorder in a subject comprising: administering to thesubject a therapeutically effective amount of a compound disclosedherein. In one embodiment, the fibrotic disorder is liver fibrosis, lungfibrosis, or kidney fibrosis.

The present invention also relates to a method of treating glaucoma orregulating intraocular pressure in a subject comprising administering tothe subject a therapeutically effective amount of a compound disclosedherein. In one aspect, the glaucoma is primary open-angle glaucoma,acute angle-closure glaucoma, pigmentary glaucoma, congenital glaucoma,normal tension glaucoma, or secondary glaucoma.

In some embodiments, the present invention is directed to a method oftreating a neoplastic disease in a subject comprising: administering tothe subject a therapeutically effective amount of a compound disclosedherein. In certain aspects, the neoplastic disorder is a lymphoma,carcinoma, leukemia, sarcoma, or blastoma. In other aspects, theneoplastic disorder is squamous cell cancer, small-cell king cancer,pituitary cancer, esophageal cancer, astrocytoma, soft tissue sarcoma,non-small cell lung cancer, adenocarcinoma of the king, squamouscarcinoma of the king, cancer of the peritoneum, hepatocellular cancer,gastrointestinal cancer, pancreatic cancer, glioblastoma, cervicalcancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breastcancer, colon cancer, colorectal cancer, endometrial or uterinecarcinoma, salivary gland carcinoma, kidney cancer, liver cancer,prostate cancer, vulval cancer, thyroid cancer, hepatic carcinoma, braincancer, endometrial cancer, testis cancer, cholangiocarcinoma,gallbladder carcinoma, gastric cancer, melanoma, or head and neckcancer.

In yet other embodiments, the present invention provides a method oftreating metabolic syndrome, insulin resistance, hyperinsulinemia, type2 diabetes, or glucose intolerance in a subject comprising administeringto the subject a therapeutically effective amount of a compounddisclosed herein.

In one embodiment, the present invention relates to a method of treatingosteoporosis or promoting bone formation in a subject comprisingadministering to the subject a therapeutically effective amount of acompound disclosed herein.

In another embodiment, the present invention relates to a method oftreating an ocular disorder having an angiogenic component comprisingadministering to the subject a therapeutically effective amount of acompound disclosed herein and an angiogenesis inhibitor. In certainaspects, the ocular disorder is age related macular degeneration (AMD),choroidal neovascularization (CNV), diabetic macular edema (DME), irisneovascularization, uveitis, neo vascular glaucoma, or retinitis ofprematurity (ROP).

DETAILED DESCRIPTION OF THE INVENTION

In the following description, and for the purposes of explanation,numerous specific details are set forth in order to provide a thoroughunderstanding of the various aspects of the invention. It will beunderstood, however, by those skilled in the relevant arts, that thepresent invention may be practiced without these specific details. Inother instances, known structures and devices are shown or discussedmore generally in order to avoid obscuring the invention. In many cases,a description of the operation is sufficient to enable one to implementthe various forms of the invention, particularly when the operation isto be implemented in software. It should be noted that there are manydifferent and alternative configurations, devices and technologies towhich the disclosed inventions may be applied. The full scope of theinventions is not limited to the examples that are described below.

It will be understood that “substituted”, “substitution” or “substitutedwith” includes the implicit proviso that such substitution is inaccordance with permitted valence of the substituted atom and thesubstituent, and that the substitution results in a stable compound,e.g., which does not spontaneously undergo transformation such as byrearrangement, cyclization, elimination, etc.

As used herein, the term “substituted” is contemplated to include allpermissible substituents of organic compounds. In a broad aspect, thepermissible substituents include acyclic and cyclic, branched andunbranched, carbocyclic and heterocyclic, aromatic and non-aromaticsubstituents of organic compounds. Illustrative substituents include,for example, those described herein below.

The term “alkyl,” as used herein unless otherwise defined, refers to astraight or branched saturated group derived from the removal of ahydrogen atom from an alkane. Representative straight chain alkyl groupsinclude -methyl, -ethyl, -n-propyl, -n-butyl, -n-pentyl, and n-heptyl.

Representative branched alkyl groups include -isopropyl, -sec-butyl,-isobutyl, -tert-butyl, -isopentyl, -neopentyl, 1-methylbutyl,2-methylbutyl, 3-methylbutyl, 1,1-dimethylpropyl and 1,2-dimethylpropyl.

As used herein, halo groups include any halogen. Examples include butare not limited to —F, —Cl, —Br, or —I.

A C₁-C₆ alkyl group includes any straight or branched, saturated orunsaturated, substituted or unsubstituted hydrocarbon comprised ofbetween one and six carbon atoms. Examples of —C₁-C₆ alkyl groupsinclude, but are not limited to methyl, ethyl, propyl, isopropyl, butyl,sec-butyl, tert-butyl, pentyl, isopentyl, neopentyl, hexyl, isohexyl,neohexyl, ethylenyl, propylenyl, 1-butenyl, 2-butenyl, 1-pentenyl,2-pentenyl, 1-hexenyl, 2-hexenyl, 3-hexenyl, acetylenyl, pentynyl,1-butynyl, 2-butynyl, 1-pentynyl, 2-pentynyl, 1-hexynyl, 2-hexynyl and3-hexynyl groups. Substituted —C₁-C₆ alkyl groups may include anyapplicable chemical moieties.

Examples of groups that may be substituted onto any of the above listed—C₁-C₆ alkyl groups include but are not limited to the followingexamples: halo, —C₁-C₆ alkyl, —O—(C₁-C₆ alkyl), C₃-C₇ cycloalkyl, —OH,—CN, —COOR′, —OC(O)R′, —NHR′, N(R′)₂, —NHC(O)R′ or —C(O)NHR′ groups. Thegroups denoted R′ above may be —H, any —C₁-C₆ alkyl, or two R′ may,optionally with a nitrogen or an oxygen atom which they are bound to,form a 3-, 4-, 5-, 6-, 7-membered ring system when the substitution is—N(R′)₂; An aryl group includes any unsubstituted or substituted phenylor napthyl group. Examples of groups that may be substituted onto ayaryl group include, but are not limited to: halo, —C₁-C₆ alkyl,—O—(C₁-C₆ alkyl), —OH, —CN, —COOR′, —OC(O)R′, NHR′, N(R′) 2, —NHC(O),R′, or —C(O)NEtR′. The group denoted R′ may be —H or any —C₁-C₆ alkyl.

A C₃-C₇ cycloalkyl group includes any 3-, 4-, 5-, 6-, or 7-memberedsubstituted or unsubstituted non-aromatic carbocyclic ring. Examples ofC₃-C₇ cycloalkyl groups include, but are not limited to, cyclopropyl,cyclobutyl, cyclopentyl, cyclopentadienyl, cyclohexyl, cyclohexenyl,cycloheptyl, cycloheptanyl, 1,3-cyclohexadienyl,-1,4-cyclohexadienyl,-1, 3-cycloheptadienyl, and-1,3, 5-cycloheptatrienylgroups. Examples of groups that may be substituted onto C₃-C₇ cycloalkylgroups include, but are not limited to: -halo, —C₁-C₆ alkyl, —O—(C₁-C₆alkyl), —OH, —CN, —COOR′, —OC(O) R′, NHR′, N(R′)2, —NHC(O)R′ or—C(O)NHR′ groups. The groups denoted R′ above include an —H or anyunsubstituted —C₁-C₆ alkyl, examples of which are listed above. Halogroups include any halogen. Examples include but are not limited to —F,—Cl, —Br, or —I.

A heterocycle may be any optionally substituted saturated, unsaturatedor aromatic cyclic moiety wherein said cyclic moiety is interrupted byat least one heteroatom selected from oxygen (O), sulfur (S) or nitrogen(N). Heterocycles may be monocyclic or polycyclic rings. For example,suitable substituents include halogen, halogenated C₁₋₆ alkyl,halogenated C₁₋₆ alkoxy, amino, amidino, amido, azido, cyano, guanidino,hydroxyl, nitro, nitroso, urea, OS(O)₂R; OS(O)₂OR, S(O)₂OR S(O)₀₋₂R,C(O)OR wherein R may be H, C₁-C₆ alkyl, aryl or 3 to 10 memberedheterocycle) OP(O)OR₁OR₂, P(O)OR₁OR₂, SO₂NR₁R₂, NR₁SO₂R₂ C(R₁)NR₂C(R₁)NOR₂, R₁ and R₂ may be independently H, C₁-C₆ alkyl, aryl or 3 to10 membered heterocycle), NR₁C(O)R₂, NR₁C(O)OR₂, NR₃C(O)NR₂R₁,C(O)NR₁R₂, OC(O)NR₁R₂. For these groups, R₁, R₂ and R₃ are eachindependently selected from H, C₁-C₆ alkyl, aryl or 3 to 10 memberedheterocycle or R₁ and R₂ are taken together with the atoms to which theyare attached to form a 3 to 10 membered heterocycle.

Possible substituents of heterocycle groups include halogen (Br, Cl, Ior F), cyano, nitro, oxo, amino, C₁₋₄ alkyl (e.g., CH₃, C₂H₅, isopropyl)C₁₋₄ alkoxy (e.g., OCH₃, OC₂H₅), halogenated C₁₋₄ alkyl (e.g., CF₃,CHF₂), halogenated C₁₋₄ alkoxy (e.g., OCF₃, OC₂F₅), COOH, COO—C₁₋₄alkyl, CO—C₁₋₄ alkyl, C₁₋₄ alkyl —S— (e.g., CH₃S, C₂H₅S), halogenatedC₁₋₄ alkyl —S— (e.g., CF₃S, C₂F₅S), benzyloxy, and pyrazolyl.

Examples of heterocycles include but are not limited to azepinyl,aziridinyl, azetyl, azetidinyl, diazepinyl, dithiadiazinyl,dioxazepinyl, dioxolanyl, dithiazolyl, furanyl, isooxazolyl,isothiazolyl, imidazolyl, morpholinyl, morpholino, oxetanyl,oxadiazolyl, oxiranyl, oxazinyl, oxazolyl, piperazinyl, pyrazinyl,pyridazinyl, pyrimidinyl, piperidyl, piperidino, pyridyl, pyranyl,pyrazolyl, pyrrolyl, pyrrolidinyl, thiatriazolyl, tetrazolyl,thiadiazolyl, triazolyl, thiazolyl, thienyl, tetrazinyl, thiadiazinyl,triazinyl, thiazinyl, thiopyranyl furoisoxazolyl, imidazothiazolyl,thienoisothiazolyl, thienothiazolyl, imidazopyrazolyl,cyclopentapyrazolyl, pyrrolopyrrolyl, thienothienyl,thiadiazolopyrimidinyl, thiazolothiazinyl, thiazolopyrimidinyl,thiazolopyridinyl, oxazolopyrimidinyl, oxazolopyridyl, benzoxazolyl,benzisothiazolyl, benzothiazolyl, imidazopyrazinyl, purinyl,pyrazolopyrimidinyl, imidazopyridinyl, benzimidazolyl, indazolyl,benzoxathiolyl, benzodioxolyl, benzodithiolyl, indolizinyl, indolinyl,isoindolinyl, furopyrimidinyl, furopyridyl, benzofuranyl,isobenzofuranyl, thienopyrimidinyl, thienopyridyl, benzothienyl,cyclopentaoxazinyl, cyclopentafuranyl, benzoxazinyl, benzothiazinyl,quinazolinyl, naphthyridinyl, quinolinyl, isoquinolinyl, benzopyranyl,pyridopyridazinyl and pyridopyrimidinyl groups.

The invention further encompasses any other physiochemical orsterochemical form that the compound may assume. Such forms includediastereomers, racemates, isolated enantiomers, hydrated forms, solvatedforms, any known or yet to be disclosed crystalline or amorphous formincluding all polymorphic crystalline forms. Amorphous forms lack adistinguishable crystal lattice and therefore lack an orderlyarrangement of structural units. Many pharmaceutical compounds haveamorphous forms. Methods of generating such chemical forms will be wellknown by one with skill in the art.

Another aspect of the invention is that the carbon atom bearing R₁ and-QW in Formula I and may have “S” or “R” configuration. Alldiastereomers, racemates, and isolated enantiomers are within the scopeof the invention.

Racemates, individual enantiomers, or diasteromers of the compound maybe prepared by specific synthesis or resolution through any method nowknown or yet to be disclosed. For example, the compound may be resolvedinto it enantiomers by the formation of diasteromeric pairs through saltformation using an optically active acid. Enantiomers are fractionallycrystallized and the free base regenerated. In another example,enantiomers may be separated by chromatography. Such chromatography maybe any appropriate method now known or yet to be disclosed that isappropriate to separate enantiomers such as HPLC on a chiral column.

The benzamide and pyrazole moiety and its intermediates may exist indifferent tautomeric forms. Tautomers include any structural isomers ofdifferent energies that have a low energy barrier to interconversion.One example is proton tautomers (prototropic tautomers.) In thisexample, the interconversions occur via the migration of a proton.Examples of prototropic tautomers include, but are not limited toketo-enol and imine-enamine isomerizations. In another exampleillustrated graphically below, proton migration between the 1-position,2-amino and 3-position nitrogen atoms of a 2-aminobenzimidazole ring mayoccur. As a result, Formulas IIa, IIb and IIc are tautomeric forms ofeach other:

Similarly, Formulas IIIa and IIIb are tautomeric forms of each other:

In some aspects of the invention the compound is in the form of apharmaceutically acceptable salt. Pharmaceutically acceptable saltsinclude any salt derived from an organic or inorganic acid. Examples ofsuch salts include but are not limited to the following: salts ofhydrobromic acid, hydrochloric acid, nitric acid, phosphoric acid andsulphuric acid. Organic acid addition salts include, for example, saltsof acetic acid, benzenesulphonic acid, benzoic acid, camphorsulphonicacid, citric acid, 2-(4-chlorophenoxy)-2-methylpropionic acid, 1,2-ethanedisulphonic acid, ethanesulphonic acid,ethylenediaminetetraacetic acid (EDTA), fumaric acid, glucoheptonicacid, gluconic acid, glutamic acid, N-glycolylarsanilic acid,4-hexylresorcinol, hippuric acid, 2-(4-hydroxybenzoyl) benzoicacid,1-hydroxy-2-naphthoicacid, 3-hydroxy-2-naphthoic acid,2-hydroxyethanesulphonic acid, lactobionic acid, n-dodecyl sulphuricacid, maleic acid, malic acid, mandelic acid, methanesulphonic acid,methyl sulpuric acid, mucic acid, 2-naphthalenesulphonic acid, pamoicacid, pantothenic acid, phosphanilic acid ((4-aminophenyl) phosphonicacid), picric acid, salicylic acid, stearic acid, succinic acid, tannicacid, tartaric acid, terephthalic acid, p-toluenesulphonic acid,10-undecenoic acid or any other such acid now known or yet to bedisclosed. It will be appreciated that such salts, provided that theyare pharmaceutically acceptable, may be used in therapy. Such salts maybe prepared by reacting the compound with a suitable acid in a mannerknown by those skilled in the art.

In some embodiments, the compounds of the present invention cause kinaseinhibition in vitro and/or in vivo. Methods of determining kinaseinhibition are well known in the art. For example, kinase activity of anenzyme and the inhibitory capacity of a test compound can be determinedby measuring enzyme specific phosphorylation of a substrate. Commercialassays and kits can be employed. For example, kinase inhibition can bedetermined using an IMAP® assay (Molecular Devices). This assay methodinvolves the use of a fluorescently-tagged peptide substrate.Phosphorylation of the tagged peptide by a kinase of interest promotesbinding of the peptide to a trivalent metal-based nanoparticle via thespecific, high affinity interaction between the phospho-group and thetrivalent metal. Proximity to the nanoparticle results in increasedfluorescence polarization. Inhibition of the kinase by a kinaseinhibitor prevents phosphorylation of the substrate and thereby limitsbinding of the fluorescently-tagged substrate to the nanoparticle. Suchan assay can be compatible with a microwell assay format, allowingsimultaneous determination of IC₅₀ of multiple compounds.

In another aspect of the present invention there is provided a method oftreating a patient suffering from a disease comprising administering toa patient in need of such treatment a therapeutically effective amountof a compound of the present invention. The phrase“therapeutically-effective amount” as used herein means that amount of acompound, material, or composition comprising a compound of the presentinvention which is effective for producing some desired therapeuticeffect in at least a sub-population of cells in an animal at areasonable benefit/risk ratio applicable to any medical treatment, e.g.,reasonable side effects applicable to any medical treatment.

Compounds of the invention are useful for treatment of patientssuffering from cardiovascular and non-cardiovascular diseases, such ashypertension, pulmonary hypertension, atherosclerosis, restenosis,coronary heart disease, cardiac hypertrophy, ocular hypertension,retinopathy, ischemic diseases, cerebral ischemia, cerebral vasospasm,penile erectile dysfunction, peripheral circulatory disorder, peripheralartery occlusive disease, glaucoma, (e.g., regulating intraocularpressure), fibroid lung, fibroid liver, fibroid kidney, chronicobstructive pulmonary disease (COPD), adult respiratory distresssyndrome, central nervous system disorders such as neuronal degenerationand spinal cord injury. Further, the compounds of the invention can beused to treat arterial thrombotic disorders such as platelet aggregationand leukocyte aggregation, and bone resorption.

In an embodiment of the invention, compounds are used to treat cerebralcavernous malformation (CCM). CCMs are vascular lesions consisting ofclusters of leaky, dilated capillaries and are associated with centralnervous system (CNS) disorders, including seizures and stroke. The lossof vascular integrity is thought to involve activation of RhoA andactivation of ROCK, leading to changes in cytoskeletal stability andincreased vascular permeability. The compounds of the invention inhibitROCK activation and restore vascular endothelial function.

The compounds of the invention can also be used to treat glaucoma. Thereare several types of glaucoma which can be treated, including, withoutlimitation, the following types. The two most common, primary open-angleglaucoma and acute angle-closure glaucoma are characterized by highocular pressure. Pigmentary glaucoma and congenital glaucoma also arecharacterized by reduced fluid outflow and high intraocular pressure(IOP). Normal tension glaucoma is thought to be due to anothermechanism, in particular poor blood flow to the optic nerve. Secondaryglaucoma can result from injury, infection, inflammation, tumor orcataracts, and is also associated with prolonged use of steroids,systemic hypertension, diabetic retinopathy, and central retinal veinocclusion.

In certain embodiments, the compounds of the invention are used to treatinflammation, including, but not limited to asthma, cardiovascularinflammation, renal inflammation, atherosclerosis and arteriosclerosis.

In some embodiments, the compounds of the invention inhibit tumor cellgrowth and metastasis, and angiogenesis, and are useful for treatingneoplastic diseases. Neoplastic diseases include any malignant growth ortumor caused by abnormal or uncontrolled cell division, and may spreadto other parts of the body through the lymphatic system or the bloodstream Neoplastic disease includes, without limitation, lymphoma (aneoplasm of lymph tissue that is usually malignant), carcinoma (anymalignant tumor derived from epithelial tissue), leukemia (malignantneoplasm of blood-forming tissues; characterized by abnormalproliferation of leukocytes), sarcoma (a usually malignant tumor arisingfrom connective tissue (bone or muscle etc.), and blastoma (malignancyin precursor cells). Non-limiting examples include squamous cell cancer,small-cell lung cancer, pituitary cancer, esophageal cancer,astrocytoma, soft tissue sarcoma, non-small cell king cancer,adenocarcinoma of the lung, squamous carcinoma of the king, cancer ofthe peritoneum, hepatocellular cancer, gastrointestinal cancer,pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, livercancer, bladder cancer, hepatoma, breast cancer, colon cancer,colorectal cancer, endometrial or uterine carcinoma, salivary glandcarcinoma, kidney cancer, liver cancer, prostate cancer, vulval cancer,thyroid cancer, hepatic carcinoma, brain cancer, endometrial cancer,testis cancer, cholangiocarcinoma, gallbladder carcinoma, gastriccancer, melanoma, and various types of head and neck cancer.

According to one aspect of the invention, the inventive compounds areused to effect weight loss and/or limit weight gain. In a preferredembodiment, the compound is ROCK2 selective. ROCK-2 inhibitors promoteweight loss in normal subjects, and limit weight gain in subjects proneto obesity.

In an embodiment of the invention, the inventive compound is used toreduce or prevent insulin resistance or restore insulin sensitivity.Accordingly, in one embodiment, the compounds of the invention are usedto promote or restore insulin-dependent glucose uptake. In anotherembodiment of the invention, a compound of the invention is used topromote or restore glucose tolerance. In another embodiment of theinvention, a compound of the invention is used to treat metabolicsyndrome. In another embodiment, a compound of the invention is used toreduce or prevent hyperinsulinemia. In an embodiment of the invention,an inventive compound is used to treat diabetes (particularly type 2diabetes). Compounds of the invention may also be used to promote orrestore insulin-mediated relaxation of vascular smooth muscle cells(VSMCs).

The invention provides methods and compounds for treating diseases anddisorders with an angiogenic component. According to the invention, incertain embodiments, such diseases and disorders are treated byadministering to a subject an effective amount of a rho kinaseinhibitor. In certain embodiments, the inventive compound is a ROCK2selective inhibitor. According to the invention, such diseases anddisorders can also be treated by administering an effective amount of arho kinase inhibitor that inhibits ROCK2, and may be ROCK2 selective,and an effective amount of an angiogenesis inhibitor. According to theinvention, ocular diseases and disorders having an angiogenic componentare treated in this manner. In one embodiment, the invention provides amethod of treating age related macular degeneration (AMD), which occursin “dry” and “wet” forms. The “wet” form of AMD causes vision loss dueto abnormal blood vessel growth (neovascularization). Bleeding, leaking,and scarring from these retinal blood vessels eventually causesirreversible damage to the photoreceptors. The dry form results fromatrophy of the retinal pigment epithelial layer, which causes visionloss through loss of photoreceptors (rods and cones) in the central partof the eye. In another embodiment, the invention provides a method oftreating choroidal neovascularization (CNV). Choroidalneovascularization is a process in which new blood vessels grow in thechoroid, through the Bruch membrane and invade the subretinal space, andis a symptom of, among other causes, age-related macular degeneration,myopia and ocular trauma. In another embodiment, the invention providesa method of treating diabetic macular edema (DME). In anotherembodiment, the invention provides a method of treating macular edemathat is secondary to branch retinal vein occlusion (BRVO) or centralretinal vein occlusion (CRVO). In other embodiments, the diseases to betreated include, without limitation, retinal neovascularization,infectious and non-infectious, corneal neovascularization infectious andnon-infectious, iris neovascularization, uveitis, neovascular glaucoma,and retinitis of prematurity (ROP). The method of treatment can beprophylactic, such as to stave off corneal neovascularization aftercorneal transplant, or to modulate the wound healing process intrabeculectomy surgery. These diseases and disorders may becharacterized as having an angiogenic component. According to theinvention, such disorders are treated by administering an inventivecompound and an angiogenesis inhibitor.

Accordingly, in one such embodiment, the disease or disorder is AMD, anda subject in need of treatment for AMD is administered an amount of aninventive compound to treat AMD. In another embodiment, the subject isadministered an inventive compound and an angiogenesis inhibitor inamounts effective to treat AMD. In such embodiments, a ROCK2-selectiveinhibitor may be preferred. In some embodiments, the angiogenesisinhibitor is a VEGFR2 antagonist. In certain such embodiments, theVEGFR2 antagonist binds to VEGF. In other such embodiments, the VEGFR2antagonist binds to VEGFR2. Such VEGFR2-binding inhibitors includeagents that bind to the extracellular domain of VEGFR2, including butnot limited to antibodies and VEGFR2-binding fragments thereof andagents that interact with the intracellular domain of VEGFR2 and blockactivation of VEGFR2-dependent signal transduction. VEGFR2 antagonistsfurther include agents that interact with other cellular components toblock VEGFR2-dependent signal transduction. In other embodiments of theinvention, other ocular diseases and disorders having an angiogeniccomponent, such as are indicated above, are similarly treated.

According to the invention, an inventive compound and an angiogenesisinhibitor are administered to a subject in amounts effective amount totreat or preventing a pathologic condition characterized by excessiveangiogenesis. Such conditions, involving for example, vascularizationand/or inflammation, include atherosclerosis, rheumatoid arthritis (RA),hemangiomas, angiofibromas, and psoriasis. Other non-limiting examplesof angiogenic disease are retinopathy of prematurity (retrolentalfibroplastic), corneal graft rejection, corneal neovascularizationrelated to complications of refractive surgery, cornealneovascularization related to contact lens complications, cornealneovascularization related to pterygium and recurrent pterygium, cornealulcer disease, and non-specific ocular surface disease,insulin-dependent diabetes mellitus, multiple sclerosis, myastheniagravis, Crohn's disease, autoimmune nephritis, primary biliarycirrhosis, acute pancreatitis, allograph rejection, allergicinflammation, contact dermatitis and delayed hypersensitivity reactions,inflammatory bowel disease, septic shock, osteoporosis, osteoarthritis,cognition defects induced by neuronal inflammation, Osier-Webersyndrome, restenosis, and fungal, parasitic and viral infections,including cytomegalovirus infections.

The invention further encompasses pharmaceutical compositions thatinclude the disclosed compound as an ingredient. Such pharmaceuticalcompositions may take any physical form necessary depending on a numberof factors including the desired method of administration and thephysicochemical and stereochemical form taken by the disclosed compoundor pharmaceutically acceptable salts of the compound. Such physicalforms include a solid, liquid, gas, sol, gel, aerosol, or any otherphysical form now known or yet to be disclosed. The concept of apharmaceutical composition including the disclosed compound alsoencompasses the disclosed compound or a pharmaceutically acceptable saltthereof without any other additive. The physical form of the inventionmay affect the route of administration and one skilled in the art wouldknow to choose a route of administration that takes into considerationboth the physical form of the compound and the disorder to be treated.Pharmaceutical compositions that include the disclosed compound may beprepared using methodology well known in the pharmaceutical art. Apharmaceutical composition that includes the disclosed compound mayinclude a second effective compound of a distinct chemical formula fromthe disclosed compound. This second effective compound may have the sameor a similar molecular target as the target or it may act upstream ordownstream of the molecular target of the disclosed compound with regardto one or more biochemical pathways.

Pharmaceutical compositions including the disclosed compound includematerials capable of modifying the physical form of a dosage unit. Inone nonlimiting example, the composition includes a material that formsa coating that holds in the compound. Materials that may be used in sucha coating, include, for example, sugar, shellac, gelatin, or any otherinert coating agent.

Pharmaceutical compositions including the disclosed compound may beprepared as a gas or aerosol. Aerosols encompass a variety of systemsincluding colloids and pressurized packages. Delivery of a compositionin this form may include propulsion of a pharmaceutical compositionincluding the disclosed compound through use of liquefied gas or othercompressed gas or by a suitable pump system. Aerosols may be deliveredin single phase, bi-phasic, or tri-phasic systems.

In some aspects of the invention, the pharmaceutical compositionincluding the disclosed compound is in the form of a solvate. Suchsolvates are produced by the dissolution of the disclosed compound in apharmaceutically acceptable solvent. Pharmaceutically acceptablesolvents include any mixtures of more than one solvent. Such solventsmay include pyridine, chloroform, propan-1-ol, ethyl oleate, ethyllactate, ethylene oxide, water, ethanol, and any other solvent thatdelivers a sufficient quantity of the disclosed compound to treat theaffliction without serious complications arising from the use of thesolvent in patients.

Pharmaceutical compositions that include the disclosed compound may alsoinclude a pharmaceutically acceptable carrier. Carriers include anysubstance that may be administered with the disclosed compound with theintended purpose of facilitating, assisting, or helping theadministration or other delivery of the compound. Carriers include anyliquid, solid, semisolid, gel, aerosol or anything else that may becombined with the disclosed compound to aid in its administration.Examples include diluents, adjuvants, excipients, water, oils (includingpetroleum, animal, vegetable or synthetic oils,) Such carriers includeparticulates such as a tablet or powder, liquids such as an oral syrupor injectable liquid, and inhalable aerosols. Further examples includesaline, gum acacia, gelatin, starch paste, talc, keratin, colloidalsilica, and urea. Such carriers may further include binders such asethyl cellulose, carboxymethylcellulose, microcrystalline cellulose, orgelatin; excipients such as starch, lactose or dextrins; disintegratingagents such as alginic acid, sodium alginate, Primogel, and corn starch;lubricants such as magnesium stearate or Sterotex; glidants such ascolloidal silicon dioxide; sweetening agents such as sucrose orsaccharin, a flavoring agent such as peppermint, methyl salicylate ororange flavoring, or coloring agents. Further examples of carriersinclude polyethylene glycol, cyclodextrin, oils, or any other similarliquid carrier that may be formulated into a capsule. Still furtherexamples of carriers include sterile diluents such as water forinjection, saline solution, physiological saline, Ringer's solution,isotonic sodium chloride, fixed oils such as synthetic mono ordiglycerides, polyethylene glycols, glycerin, cyclodextrin, propyleneglycol or other solvents; antibacterial agents such as benzyl alcohol ormethyl paraben; antioxidants such as ascorbic acid or sodium bisulfite;chelating agents such as ethylenediaminetetraacetic acid; buffers suchas acetates, citrates or phosphates and agents for the adjustment oftonicity such as sodium chloride or dextrose, thickening agents,lubricating agents, and coloring agents.

The pharmaceutical composition including the disclosed compound may takeany of a number of formulations depending on the physicochemical form ofthe composition and the type of administration. Such forms includesolutions, suspensions, emulsions, tablets, pills, pellets, capsules,capsules including liquids, powders, sustained-release formulations,directed release formulations, lyophylates, suppositories, emulsions,aerosols, sprays, granules, powders, syrups, elixirs, or any otherformulation now known or yet to be disclosed. Additional examples ofsuitable pharmaceutical carriers are described in “Remington'sPharmaceutical Sciences” by E. W. Martin, hereby incorporated byreference in its entirety.

Methods of administration include, but are not limited to, oraladministration and parenteral administration. Parenteral administrationincludes, but is not limited to intradermal, intramuscular,intraperitoneal, intravenous, subcutaneous, intranasal, epidural,sublingual, intranasal, intracerebral, intraventricular, intrathecal,intravaginal, transdermal, rectal, by inhalation, or topically to theears, nose, eyes, or skin. Other methods of administration include butare not limited to infusion techniques including infusion or bolusinjection, by absorption through epithelial or mucocutaneous liningssuch as oral mucosa, rectal and intestinal mucosa. Compositions forparenteral administration may be enclosed in ampoule, a disposablesyringe or a multiple-dose vial made of glass, plastic or othermaterial.

Administration may be systemic or local. Local administration isadministration of the disclosed compound to the area in need oftreatment. Examples include local infusion during surgery; topicalapplication, by local injection; by a catheter; by a suppository; or byan implant. Administration may be by direct injection at the site (orformer site) of a cancer, tumor, or precancerous tissue or into thecentral nervous system by any suitable route, including intraventricularand intrathecal injection. Intraventricular injection may be facilitatedby an intraventricular catheter, for example, attached to a reservoir,such as an Ommaya reservoir. Pulmonary administration may be achieved byany of a number of methods known in the art. Examples include use of aninhaler or nebulizer, formulation with an aerosolizing agent, or viaperfusion in a fluorocarbon or synthetic pulmonary surfactant. Thedisclosed compound may be delivered in the context of a vesicle such asa liposome or any other natural or synthetic vesicle.

A pharmaceutical composition formulated so as to be administered byinjection may be prepared by dissolving the disclosed compound withwater so as to form a solution. In addition, a surfactant may be addedto facilitate the formation of a homogeneous solution or suspension.Surfactants include any complex capable of non-covalent interaction withthe disclosed compound so as to facilitate dissolution or homogeneoussuspension of the compound.

Pharmaceutical compositions including the disclosed compound may beprepared in a form that facilitates topical or transdermaladministration. Such preparations may be in the form of a solution,emulsion, ointment, gel base, transdermal patch or iontophoresis device.Examples of bases used in such compositions include opetrolatum,lanolin, polyethylene glycols, beeswax, mineral oil, diluents such aswater and alcohol, and emulsifiers and stabilizers, thickening agents,or any other suitable base now known or yet to be disclosed.

Examples that represent different aspects of the invention follow. Suchexamples should not be construed as limiting the scope of thedisclosure. Alternative mechanistic pathways and analogous structureswithin the scope of the invention would be apparent to those skilled inthe art.

Elements and acts in the examples are intended to illustrate theinvention for the sake of simplicity and have not necessarily beenrendered according to any particular sequence or embodiment.

EXAMPLES Example 1. Synthesis of Kinase Inhibitors

Different aspects of the invention may be prepared via the generalsynthetic procedures outlined below. It will be apparent to one skilledin the art how to prepare the other aspects of the invention by choiceof proper and relevant starting materials, synthetic intermediates andreagents.

A compound of Formula I can be prepared according to Scheme 1. A1-[bis(dimethylamino)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium3-oxid hexafluorophosphate (HATU) mediated amide coupling reactionbetween carboxylic acid 1 and amine 2 in DMF (N,N-dimethylformamide)generates structure 3. Under the same conditions, amine 4 reacts with 5to generate structure 6.

Another compound of Formula I can be prepared according to Scheme 2. Thecoupling of amine 4 and phenyl chloroformate yields structure 7, whichis treated with amine 2 to afford urea 9.

A compound of Formula V can be prepared according to Scheme 3. Anilinediamine 10 and isothiocyanate 11 are heated in THF and the cyclizationof the resulting thiourea with iodomethane yields aminoimidazole 12.

Example 2. Kinase Inhibitor Compounds

Non-limiting examples illustrative of the invention include those shownin Table 1.

TABLE 1 Non-limiting examples of kinase inhibitor compounds. Unlessspecified, example compounds with a chiral center represent racemicmixture of the corresponding R and S enantiomers and all racemates andisolated enantiomers are within the scope of the invention. ID StructureM + 1 1

327 2

327 3

310 4

312 5

324 6

342 7

338 8

326 9

368 10

356 11

395 12

338 13

309 14

327 15

327 16

343 17

345 18

337 19

336 20

355 21

357 22

327 23

341 24

341 25

324 26

312 27

354 28

338 29

368 30

324 31

338 32

320 33

350 34

361 35

331 36

338 37

343 38

340 39

376 40

365 41

390 42

394 43

406 44

353 45

447 46

436 47

342 48

342 49

342 50

342 51

368 52

338 53

352 54

354 55

354 56

352 57

352

Example 3. Synthesis of Compound 14

Step 1

methyl2-(3-fluorophenyl)-2-(3-(pyridin-4-yl)-1H-pyrazole-5-carboxamido)acetate

To a mixture of 3-(pyridin-4-yl)-1H-pyrazole-5-carboxylic acid (95 mg,0.50 mmol), methyl 2-amino-2-(3-fluorophenyl)acetate HCl salt (143 mg,0.65 mmol) and diisopropylethylamine (0.26 mL, 1.5 mmol) in DMF wasadded HATU (248 mg, 0.65 mmol). The reaction was stirred at roomtemperature for 3 h, quenched with water and extracted with ethylacetate. The organic layer was dried, concentrated and purified byBIOTAGE® column chromatography to give methyl2-(3-fluorophenyl)-2-(3-(pyridin-4-yl)-1H-pyrazole-5-carboxamido)acetate(126 mg).

Step 2

To a solution of methyl2-(3-fluorophenyl)-2-(3-(pyridin-4-yl)-1H-pyrazole-5-carboxamido)acetate (62 mg, 0.17 mmol) in MeOH was added sodium borohydride (13 mg,0.34 mmol). The reaction was stirred overnight, quenched with NaOH (1N)and concentrated. The residue was purified by C-18 BIOTAGE® columnchromatography to give Compound 14 (29 mg).

Example 4. Synthesis of Compound 39

Step 1

6-chlorothieno[2,3-b]pyridine-2-carboxylic acid

To a suspension of 2-bromo-6-chlorothieno[2,3-b]pyridine (100 mg, 0.40mmol) in ether was added n-butyllithium (0.29 mL, 2.5 M, 0.72 mmol)dropwise at −40° C. The reaction was stirred for 0.5 h, quenched withexcess dry ice and partitioned between water and ethyl acetate. Theorganic layer was dried, concentrated and purified by C-18 BIOTAGE®column chromatography to give 6-chlorothieno[2,3-b]pyridine-2-carboxylicacid (36 mg).

Step 2

6-chloro-N-(3-methoxybenzyl)thieno[2,3-b]pyridine-2-carboxamide

6-chloro-N-(3-methoxybenzyl)thieno[2,3-b]pyridine-2-carboxamide wasprepared from 6-chlorothieno[2,3-b]pyridine-2-carboxylic acid byfollowing the synthesis method of Step 2 in Example 3.

Step 3

A mixture of6-chloro-N-(3-methoxybenzyl)thieno[2,3-b]pyridine-2-carboxamide (25 mg,0.075 mmol), pyridine-4-boronic acid (30 mg, 0.22 mmol),[1,1′-Bis(diphenylphosphino)ferrocene]dichloropalladium(II) (5 mg, 5%)and cesium carbonate (95 mg, 0.3 mmol) in 1,4-dioxane was heated to 100°C. and stirred under nitrogen atmosphere overnight. The mixture wasfiltered through a Celite pad. The filtrate was concentrated andpurified by BIOTAGE® C-18 column chromatography to give Compound 39 (12mg).

Example 5. ROCK1 and ROCK2 Kinase Inhibition Assays

The following assay protocol is for measuring the phosphorylation of apeptide substrate (FAM-KKLRRTLSVA-OH wherein FAM is carboxyfluorescein).The peptide is >98% purity by Capillary Electrophoresis. The peptide isphosphorylated by the protein kinase ROCK1 or ROCK2. The ROCK1 or ROCK2enzyme, substrate, and cofactors (ATP and Mg²⁺) are combined in a wellof a microtiter plate and incubated for 3 hours at 25° C. At the end ofthe incubation, the reaction is quenched by the addition of anEDTA-containing buffer. The substrate and product are separated andquantified electrophoretically using the microfluidic-based LABCHIP®3000 Drug Discovery System from Caliper Life Sciences (Hopkinton,Mass.).

The components of the assay mixture are:

-   -   100 mM HEPES, pH 7.5    -   0.1% BSA    -   0.01% Triton X-100    -   1 mM DTT    -   10 mM MgCl₂    -   10 μM Sodium Orthovanadate    -   10 μM Beta-Glycerophosphate    -   5 μM ATP (for ROCK1) or 7 μM ATP (for ROCK2)    -   1% DMSO (from compound)    -   1.25 μM FAM-KKLRRTLSVA-OH    -   3 nM ROCK1 or 2.5 nM ROCK2 enzyme

Substrate and product peptides present in each sample are separatedelectrophoretically using the LABCHIP® 3000 capillary electrophoresisinstrument. As substrate and product peptides are separated two peaks offluorescence are observed. Change in the relative fluorescence intensityof the substrate and product peaks is the parameter measured reflectingenzyme activity. Capillary electrophoregramms (RDA acquisition files)are analyzed using HTS Well Analyzer software (Caliper Life Sciences,Hopkinton, Mass.). The kinase activity in each sample is determined asthe product to sum ratio (PSR): P/(S+P), where P is the peak height ofthe product peptide and S is the peak height of the substrate peptide.For each compound, enzyme activity is measured at various concentrations(12 concentrations of compound spaced by 3× dilution intervals).Negative control samples (0%—inhibition in the absence of inhibitor) andpositive control samples (100%-inhibition in the presence of 20 mM EDTA)are assembled in replicates of four and are used to calculate%-inhibition values for each compound at each concentration. Percentinhibition (Pinh) is determined using the following equation:

Pinh=(PSR0%−PSRinh)/(PSR0%−PSR100%)*100

where PSRinh is the product sum ratio in the presence of inhibitor,PSR0% is the average product sum ratio in the absence of inhibitor, andPSR100% is the average product sum ratio in 100%-inhibition controlsamples. The IC₅₀ values of inhibitors are determined by fitting theinhibition curves (Pinh versus inhibitor concentration) by 4 parametersigmoidal dose-response model using XLfit 4 software (IBDS).

This assay can be used to test the activity of each of the exemplarycompounds identified in Table 1. It is expected that each of thesecompounds will demonstrate inhibition of the protein kinase ROCK1 and/orROCK2.

Example 6. Cell Viability Assay

Cell viability in the presence of varying concentrations of the abovelisted compounds at different time points was used to assesscytotoxicity and the effect of the compounds on cell proliferation. IC₅₀(or percent activity) data for the compounds of the present invention inK562 or MV411 cell lines are summarized in Table 2.

Cell Viability Assay—

Cell viability was measured by the CELLTITER-GlO® cell viability assayfrom Promega (Madison, Wis.). The CELLTITER-GlO® Luminescent CellViability Assay is a homogeneous method to determine the number ofviable cells in culture based on quantitation of the ATP present, whichsignals the presence of metabolically active cells. Following treatment,CELLTITER-GlO® is added to treatment wells and incubated at 37° C.luminescence values were measured at using a Molecular DevicesSpectramax microplate reader.

Experimental Design

Single Agent Studies—

Cells were grown to 70% confluency, trypsinized, counted, and seeded in96 well flat-bottom plates at a final concentration of 2.5×10³-5×10³cells/well (Day 0). Cells were allowed to incubate in growth media for24 hours. Treatment with the test agents or standard agents began on Day1 and continued for 72 hours. At the 72-hour time point, treatmentcontaining media was removed. Viable cell numbers were quantified by theCELLTITER-GLO® cell viability assay as described above. Results fromthese studies were used to calculate an IC₅₀ value (concentration ofdrug that inhibits cell growth by 50 percent of control) for eachcompound.

Data Collection—

For single agent and combination studies, data from each experiment wascollected and expressed as % Cell Growth using the followingcalculation:

% Cell Growth=(f _(test) /f _(vehicle))×100

Where f_(test) is the luminescence of the tested sample, and f_(vehicle)is the luminescence of the vehicle in which the drug is dissolved. Doseresponse graphs and IC₅₀ values were generated using Prism 6 software(GraphPad) using the following equation:

Y=(Top−Bottom)/(1+10^(((log IC50−X)−HillSlope)))

Where X is the logarithm of the concentration and Y is the response. Ystarts at the Bottom and goes to the Top with a sigmoid shape.

Example 7. ROCK1 and ROCK2 Kinase Inhibition and Cell Viability AssayResults

The protocols outlined in Examples 5 and 6 were followed to test ROCK1and ROCK2 kinase inhibition and cancer cell viability with compoundsfrom Table 1. As shown in Table 2, the compounds demonstrated inhibitionof the ROCK1 and ROCK2 kinases and growth of cancer cells.

The experiments also evaluated the selectivity of the compounds forinhibiting growth of cancer cells carrying a mutation in the Flt3 gene.The MV411 cell line expresses the mutant allele of Flt3 with internaltandem duplications (ITD) of the gene. See Quentmeier et al., “FLT3Mutations in Acute Myeloid Leukemia Cell Lines,” Leukemia 17(1), 2003,120-124. K562 is a chronic myeloid leukemia cell line that does notexpress FLT3 protein. See Grafone et al., “Monitoring of FLT3Phosphorylation Status and Its Response to Drugs By Flow Cytometry inAML Blast Cells,” Hematol Oncol. 26(3), 2008, 159-166. Patients withITD-FLT3⁺ acute myeloid leukemia (AML) experience an extremely poorprognosis. Surprisingly, many of the compounds demonstrated greaterefficacy with the MV11 cells than with the K562 cells suggesting thatthese compounds could be used to effectively treat ITD-FLT3⁺ AML.

TABLE 2 ROCK1 and ROCK2 kinase inhibition and cell viability ROCK1 ROCK2FLT3-ITD⁺ Compound IC₅₀ IC₅₀ K562* MV411* Selectivity ID (nM) (nM) (μM)(μM) (K562/MV411) 1 207 98 >100 67.6 >1 3 >100 43.7 >2 5 73 16.1 43.610.2 4 7 102 22.9 89.1 7.6 12 9 237 39.1 97.2 39.8 2 11 63.6 22.1 >10014.5 >7 25 130 28.7 85 34 3 40 65.1 17.4 >100 7.1 >14 47 17.8 7.8 2 486400 1870 49 17.8 7.8 2 50 1010 239 >100 7.8 >13 51 >100 17.0 >6 52 348.7 >100 11.2 >9 53 64.6 93.3 1 54 >100 12.6 >8 Compound 3.3 2.8 0.8 0.71 A** *MV411 is ITD-FLT3⁺, and K562 does not express ITD-FLT3.Rho-associated kinase may be manipulated for the treatment of ITD-FLT3⁺AML as reported in Onish et at., “Internal Tandem Duplication Mutationsin FLT3 Gene Augment Chemotaxis to Cxcl12 Protein by Blocking theDown-regulation of the Rho-associated Kinase via the Cxcl12/Cxcr4Signaling Axis,” J. Biol. Chem. 289 (45), 2014, 31053-31065. **CompoundA is shown below and is described by Schirok et al., “Design andSynthesis of Potent and Selective Azaindole-Based Rho Kinase (ROCK)Inhibitors,” ChemMedChem 3, 2008, 1893-1904.

Compound A

Unless defined otherwise, all technical and scientific terms herein havethe same meaning as commonly understood by one of ordinary skill in theart to which this invention belongs. All publications, patents, andpatent publications cited are incorporated by reference herein in theirentirety for all purposes.

It is understood that the disclosed invention is not limited to theparticular methodology, protocols and materials described as these canvary. It is also understood that the terminology used herein is for thepurposes of describing particular embodiments only and is not intendedto limit the scope of the present invention which will be limited onlyby the appended claims.

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed by the following claims.

1. A compound of Formula (I):

or an enantiomer, a mixture of enantiomers, or a mixture of two or morediastereomers thereof; or a pharmaceutically acceptable salt, solvate,or hydrate thereof; wherein: A is indazol-3-yl, pyrazol-4-yl,

wherein (i) G is CR′ or N; (ii) X is hydrogen, C₁₋₆ alkyl, C₃₋₇cycloalkyl, —OR₂ or —NR₃R₄; and (iii) R′, R″, R₂, R₃ and R₄ areindependently —H, C₁₋₆ alkyl or C₃₋₇ cycloalkyl; Z is

wherein G₁, G₂ and G₃ are independently CH or N; Z′ is a bond; R is —H,C₁₋₆ alkyl or C₃₋₇ cycloalkyl; R₁ is —H; Q is a bond or or methyl; J isa bond; W is —H or —OH, and Ar is a phenyl, naphthyl, or C₅₋₁₀heterocycle, each of which is substituted with halo, —OH, —CN,—COOR_(a), —OR_(a), —SR_(a), —OC(O)R_(a), —NHR_(a), —NR_(a)R_(b),—NHC(O)R_(a), —NHC(O)NR_(a)R_(b), —C(O)NR_(a)R_(b), —NS(O)₂R_(a),—S(O)₂NR_(a)R_(b), —S(O)₂R_(a), guanidino, nitro, nitroso, C₁₋₆ alkyl,aryl, C₃₋₇ cycloalkyl, or 3- to 10-membered heterocycle, wherein theC₁₋₆ alkyl, aryl, C₃₋₇ cycloalkyl, or 3 to 10-membered heterocycle isunsubstituted or substituted with one or more of halo, —OH, —CN,—COOR_(a), —OR_(a), —SR_(a), —OC(O)R_(a), —NHR_(a), —NR_(a)R_(b),—NHC(O)R_(a), —NHC(O)NR_(a)R_(b), —C(O)NR_(a)R_(b), —NS(O)₂R_(a),—S(O)₂NR_(a)R_(b), —S(O)₂R_(a), guanidino, nitro, nitroso, C₁₋₆ alkyl,aryl, or C₃₋₇ cycloalkyl; wherein each of R_(a) and R_(b) isindependently —H or C₁₋₆ alkyl; and optionally R_(a) and R_(b) togetherattaching to N or O form a 4- to 8-membered heterocycle.
 2. The compoundof claim 1, wherein wherein A is pyrazol-4-yl.
 3. The compound of claim2, wherein G₁ is N, and G₂ and G₃ are CH.
 4. The compound of claim 3,wherein Q is methyl.
 5. The compound of claim 4, wherein Ar is asubstituted phenyl.
 6. The compound of claim 5, wherein the phenyl issubstituted with —OR_(a), and R_(a) is —H or C₁₋₆ alkyl.
 7. The compoundof claim 51, wherein R is —H. 8-29. (canceled)
 30. A method of treatinga neoplastic disease in a subject comprising: administering to thesubject a therapeutically effective amount of a compound of claim
 1. 31.The method of claim 30, wherein the neoplastic disorder is a lymphoma,carcinoma, leukemia, sarcoma, or blastoma.
 32. The method of claim 31,wherein the neoplastic disorder is acute myeloid leukemia (AML).
 33. Themethod of claim 32, wherein the AML is ITD-FLT3⁺ AML. 34-38. (canceled)39. The compound of claim 3, wherein Ar is a substituted phenyl.
 40. Thecompound of claim 39, wherein the phenyl is substituted with —OR_(a) andR_(a) is —H or C₁₋₆ alkyl.
 41. The compound of claim 40, wherein R_(a)is methyl.
 42. The compound of claim 41, wherein R is —H.
 43. Thecompound of claim 2, wherein Ar is a substituted phenyl.
 44. Thecompound of claim 43, wherein the phenyl is substituted with —OR_(a) andR_(a) is —H or C₁₋₆ alkyl.
 45. The compound of claim 44, wherein R_(a)is methyl.
 46. The compound of claim 45, wherein R is —H.
 47. Thecompound of claim 39, wherein Q is methyl and W is —H.
 48. The compoundof claim 47, wherein the configuration of the compound is R.
 49. Thecompound of claim 39, wherein Q is methyl and W is —OH.
 50. The compoundof claim 49, wherein the configuration of the compound is S.
 51. Thecompound of claim 6, wherein R_(a) is methyl.